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Structural insights of RmXyn10A – A prebiotic-producing GH10 xylanase with a non-conserved aglycone binding region

Aronsson, Anna LU ; Güler, Fatma; Petoukhov, Maxim V.; Crennell, Susan J.; Svergun, Dmitri I.; Linares-Pastén, Javier A. LU and Nordberg Karlsson, Eva LU (2018) In Biochimica et Biophysica Acta - Proteins and Proteomics 1866(2). p.292-306
Abstract

Hydrolysis of arabinoxylan (AX) by glycoside hydrolase family 10 (GH10) xylanases produces xylo- and arabinoxylo-oligosaccharides ((A)XOS) which have shown prebiotic effects. The thermostable GH10 xylanase RmXyn10A has shown great potential to produce (A)XOS. In this study, the structure of RmXyn10A was investigated, the catalytic module by homology modelling and site-directed mutagenesis and the arrangement of its five domains by small-angle X-ray scattering (SAXS). Substrate specificity was explored in silico by manual docking and molecular dynamic simulations. It has been shown in the literature that the glycone subsites of GH10 xylanases are well conserved and our results suggest that RmXyn10A is no exception. The aglycone subsites... (More)

Hydrolysis of arabinoxylan (AX) by glycoside hydrolase family 10 (GH10) xylanases produces xylo- and arabinoxylo-oligosaccharides ((A)XOS) which have shown prebiotic effects. The thermostable GH10 xylanase RmXyn10A has shown great potential to produce (A)XOS. In this study, the structure of RmXyn10A was investigated, the catalytic module by homology modelling and site-directed mutagenesis and the arrangement of its five domains by small-angle X-ray scattering (SAXS). Substrate specificity was explored in silico by manual docking and molecular dynamic simulations. It has been shown in the literature that the glycone subsites of GH10 xylanases are well conserved and our results suggest that RmXyn10A is no exception. The aglycone subsites are less investigated, and the modelled structure of RmXyn10A suggests that loop β6α6 in the aglycone part of the active site contains a non-conserved α-helix, which blocks the otherwise conserved space of subsite +2. This structural feature has only been observed for one other GH10 xylanase. In RmXyn10A, docking revealed two alternative binding regions, one on either side of the α-helix. However, only one was able to accommodate arabinose-substitutions and the mutation study suggests that the same region is responsible for binding XOS. Several non-conserved structural features are most likely to be responsible for providing affinity for arabinose-substitutions in subsites +1 and +2. The SAXS rigid model of the modular arrangement of RmXyn10A displays the catalytic module close to the cell-anchoring domain while the carbohydrate binding modules are further away, likely explaining the observed lack of contribution of the CBMs to activity.

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author
organization
publishing date
type
Contribution to journal
publication status
published
subject
keywords
(A)XOS, Homology modelling, Manual docking, Molecular dynamics, Rhodothermus marinus, SAXS
in
Biochimica et Biophysica Acta - Proteins and Proteomics
volume
1866
issue
2
pages
15 pages
publisher
Elsevier
external identifiers
  • scopus:85034811800
ISSN
1570-9639
DOI
10.1016/j.bbapap.2017.11.006
language
English
LU publication?
yes
id
73b7f168-88bc-41c8-a185-18ddf626eb64
date added to LUP
2017-12-07 11:04:44
date last changed
2017-12-07 11:04:44
@article{73b7f168-88bc-41c8-a185-18ddf626eb64,
  abstract     = {<p>Hydrolysis of arabinoxylan (AX) by glycoside hydrolase family 10 (GH10) xylanases produces xylo- and arabinoxylo-oligosaccharides ((A)XOS) which have shown prebiotic effects. The thermostable GH10 xylanase RmXyn10A has shown great potential to produce (A)XOS. In this study, the structure of RmXyn10A was investigated, the catalytic module by homology modelling and site-directed mutagenesis and the arrangement of its five domains by small-angle X-ray scattering (SAXS). Substrate specificity was explored in silico by manual docking and molecular dynamic simulations. It has been shown in the literature that the glycone subsites of GH10 xylanases are well conserved and our results suggest that RmXyn10A is no exception. The aglycone subsites are less investigated, and the modelled structure of RmXyn10A suggests that loop β<sub>6</sub>α<sub>6</sub> in the aglycone part of the active site contains a non-conserved α-helix, which blocks the otherwise conserved space of subsite +2. This structural feature has only been observed for one other GH10 xylanase. In RmXyn10A, docking revealed two alternative binding regions, one on either side of the α-helix. However, only one was able to accommodate arabinose-substitutions and the mutation study suggests that the same region is responsible for binding XOS. Several non-conserved structural features are most likely to be responsible for providing affinity for arabinose-substitutions in subsites +1 and +2. The SAXS rigid model of the modular arrangement of RmXyn10A displays the catalytic module close to the cell-anchoring domain while the carbohydrate binding modules are further away, likely explaining the observed lack of contribution of the CBMs to activity.</p>},
  author       = {Aronsson, Anna and Güler, Fatma and Petoukhov, Maxim V. and Crennell, Susan J. and Svergun, Dmitri I. and Linares-Pastén, Javier A. and Nordberg Karlsson, Eva},
  issn         = {1570-9639},
  keyword      = {(A)XOS,Homology modelling,Manual docking,Molecular dynamics,Rhodothermus marinus,SAXS},
  language     = {eng},
  month        = {02},
  number       = {2},
  pages        = {292--306},
  publisher    = {Elsevier},
  series       = {Biochimica et Biophysica Acta - Proteins and Proteomics},
  title        = {Structural insights of RmXyn10A – A prebiotic-producing GH10 xylanase with a non-conserved aglycone binding region},
  url          = {http://dx.doi.org/10.1016/j.bbapap.2017.11.006},
  volume       = {1866},
  year         = {2018},
}